Like the cell population of tracheas in vivo, the population of cells obtained after pronase digestion of rabbit tracheas consists of ciliated, Clara-like, mucous-secreting, and basal cells. It seems likely that the nutritional requirements for cell proliferation and maintenance of these various cells types are different. It appears that the proliferation of tracheal cells in vitro is dependent on the presence of specific growth hormones and nutrients. We have shown that nutrients also play an important role in differentiation of tracheal cells in vitro: depending on whether vitamin A derivatives or serum are present, keratinization is inhibited or stimulated respectively. Also the substratum provided for trachea cells has an influence on the proliferation, attachment and maintenance of these cells: ciliated cells can be maintained for several weeks on an extracellular matrix from corneal endothelial cells, whereas ciliated cells are absent when cells are plated directly onto plastic culture dishes. Whether cell-cell interactions play a role in this process has to be determined. The objectives of our research is to understand the importance of cell-cell and cell-substratum interactions for cell proliferation, maintenance and differentiation of trachea cells. To approach these objectives we are growing cells on various substrata: different collagens and different extracellular matrices synthesized by various cell lines and determine their effect on attachment, maintenance and differentiation. Moreover, we are identifying the cell surface components and secreted products of these cells (glycoproteins, glycolipids, and proteoglycans) and try to determine the importance of the carbohydrates in cell-cell and cell-substratum interactions. Using this approach we also are attempting to establish specific markers for the various cell types.